Our long term objective is to gain a greater understanding of postreplicative DNA modifications, with respect to their nature, mechanism and specificity of occurrence and their role in gene expression. We are primarily interested in normally occurring modifications controlled by cells or viruses (viz. enzymatic processes, rather than chemically-induced modifications) since these are more likely to be of biological importance in the growth and development of the organism. Studies will be with both prokaryote and eukaryote systems. A partial list of our specific aims is as follows: (1) to determine the distribution of 5-methylcytosine (m5C) residues in the chromatin of several vertebrates, with respect to nucleosome core and linker DNA locations; (2) to study the effect of heterologous (cloned) DNA methylation on gene expression and mutation in yeast; (3) to study the effect of 5-azacytidine on gene expression and extrachromosomal DNA (plasmid) rearrangements in yeast; (4) to survey various eukaryote DNAs for methylated bases by means of HPLC and mass spectrometry [particular emphasis will be placed on certain insects and fungi where our current limit of resolution indicates less than 1 m5C per 2,000 C residues]; (5) to sequence various forms of the T4 dam methylase gene and to elucidate domains of enzymic activity and specificity; (6) to purify and characterize various forms of the T4 dam methylase enzyme activity; (7) to study regulation of the T4 dam gene following phage infection; (8) to study the regulation of the Mu mom and gin genes (with respect to the roles of the phage dad and host dam products); and (9) to determine the effect of DNA methylation on supertwisting of oriC plasmids.